Engineering mammalian cells to produce plant-specific N-glycosylation on proteins
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Protein N-glycosylation is an essential and highly conserved posttranslational modification found in all eukaryotic cells. Yeast, plants and mammalian cells, however, produce N-glycans with distinct structural features. These species-specific features not only pose challenges in selecting host cells for production of recombinant therapeutics for human medical use but also provide opportunities to explore and utilize species-specific glycosylation in design of vaccines. Here, we used reverse cross-species engineering to stably introduce plant core alpha 3fucose (alpha 3Fuc) and beta 2xylose (beta 2Xyl) N-glycosylation epitopes in the mammalian Chinese hamster ovary (CHO) cell line. We used directed knockin of plant core fucosylation and xylosylation genes (AtFucTA/AtFucTB and AtXylT) and targeted knockout of endogenous genes for core fucosylation (fut8) and elongation (B4galt1), for establishing CHO cells with plant N-glycosylation capacities. The engineering was evaluated through coexpression of two human therapeutic N-glycoproteins, erythropoietin (EPO) and an immunoglobulin G (IgG) antibody. Full conversion to the plant-type alpha 3Fuc/beta 2Xyl bi-antennary agalactosylated N-glycosylation (G0FX) was demonstrated for the IgG1 produced in CHO cells. These results demonstrate that N-glycosylation in mammalian cells is amenable for extensive cross-kingdom engineering and that engineered CHO cells may be used to produce glycoproteins with plant glycosylation.
Original language | English |
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Journal | Glycobiology |
Volume | 30 |
Issue number | 8 |
Pages (from-to) | 528-538 |
Number of pages | 11 |
ISSN | 0959-6658 |
DOIs | |
Publication status | Published - 2020 |
- core fucose, core xylose, cross-species glycoengineering, plant-type N-glycosylation, Chinese hamster ovary cells, MONOCLONAL-ANTIBODIES, O-GLYCOSYLATION, CHO-CELLS, IGG-FC, GLYCANS, FUCOSE, BIOSYNTHESIS, XYLOSE, LEWIS, ALPHA-4-FUCOSYL-TRANSFERASE
Research areas
ID: 251309969